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Grifola frondosa (Dicks.) Gray (Meripilaceae) is a big size, perennial, infrequent lignicolous basidiomycetes, traditionally known as Maitake. Brown or grayish overlapped caps, with a wavy margin, characterize the sporophore. In the wild, it is usually found at the base of living oak trees but it is also cultivated. G. frondosa is mainly distributed in Japan, Asia, Eastern America, and Europe. In Italy, is infrequent but widely distributed in all regions. Field investigation carried out in autumn 2016 in Sicily (southern Italy) permitted to observe an unusual white G. frondosa growing at the base of a living tree of Quercus pubescens Willd. s.l. Once harvested, the mushroom sample was cleaned of debris with a knife, cut into pieces and placed in plastic bags for freezing, then transported to the laboratory and preserved at -80 °C. Finally the frozen mushroom was lyophilized.The morphological studies were carried out under the microscope using KOH 3% and ammoniacal Congo Red and performed in a combination with DNA sequence analyses of the Internal Trancribed Spacer (ITS) region. The chemical and nutritional composition (ashes, water, proteins, fats, carbohydrates, vitamins, nitrogen, and organic and inorganic substances) was evaluated according to standard methodologies. The antibacterial activity against four Gram-positive and Gramnegative bacteria strains: Staphylococcus aureus ATTC 25923, S. epidermidis ATTC 12228, Pseudomonas aeruginosa ATCC 15442, and Escherichia coli ATCC 25922 was also evaluated. The minimal inhibitory concentration (MIC) of G. frondosa extracts was determinated by a micro-method by diluiting in a range from 50% to 0,6% v/v. Tryptic soy broth (TSB, Sigma) was used as media. As a result of morphological analysis the white G. frondosa showed basidiospores of slightly narrowed shape than the typical Maitake (5 - 7 x 3.2 – 4 μm). Besides, sterigmata are long up to 6 μm, sturdy, with the base of diameter up to 1.5 μm. These microscopical features are not reported in Ryvarden while Bernicchia illustrated for the typical Maitake sterigmata “thin and very divaricate”. Clamp connections are present but rare in the contest. Cystidia and cystidioles are absent. The identity of the white G.frondosa was confirmed by analysis of ITS sequence. The results of nutritional composition analysis (Table 1) showed that the white G. frondosa is richer in nutritional elements than the cultivated brown Maitake analyzed by Stamets. In particular the white Maitake possess a very high content of Ca, Fe, K, Na, and Cu. As regards vitamins only the value of vitamin D2 is similar to the value reported by other authors. The other vitamin contents are lower. The cold extract of G.frondosa has been showed to be effective in inhibiting the growth of S. epidermidis 12228 and P. aeruginosa 15442 at the maximum screening concentration of 50% v/v. In our opinion the white Maitake is a promising and potential new medicinal mushroom. Further investigations are currently in progress to evaluate potential application as cultivated and medicinal mushroom.

Antibiotic resistance of common pathogenic microorganisms is a topic of great concern that has finally received media attention and entered into the political agenda of world leaders. Drug-resistant bacteria are cause of thousands of deaths worldwide, then there is an urgent need for new antimicrobials, otherwise we risk losing the ability to control effectively the infectious diseases. Such emergence can be faced looking also at not usual source of antimicrobial agents, for example medicinal mushrooms. With the objective to tackle Gram-positive and Gram-negative pathogens, we focused on two edible desert truffles mushrooms Tirmania pinoyi and Terfezia boudieri as origin of new antimicrobial peptides (AMPs), active not only against free-living microorganisms (planktonic), but also against complex communities of pathogens (biofilms) that are intrinsically resistant to conventional antibiotics. In particular, in vitro antibacterial activity of acid-soluble protein extracts (aqueous extracts) of the two above-mentioned species were investigated against two important human pathogenic reference strains Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 15442. The acid-soluble protein extracts peptide fractions (

Staphylococcus aureus, due to its wide arsenal of virulence factors, is a very versatile pathogen responsible for a wide variety of infectious diseases. The virulence factors include the cell-wall associated proteins that have a direct role in the first stage of pathogenesis. The Sortase A (SrtA) transpeptidase is responsible for covalent anchoring to the cell wall of various surface proteins and it is considered a good target to design new antivirulence agents. In this study, we report the identification of an inhibitor of SrtA afforded from the random screening of a small molecular library of around 150 synthetic compounds, through a high throughput assay by using the standard Dabcyl-QALPETGEE-Edans fluorescence resonance energy transfer (FRET)-peptide substrate for measurement of enzyme activity. Such hit compound showed an IC50 value of 80µM. A group of derivatives of the hit compound has been obtained and evaluated for their activity as SrtA inhibitors. The efficacy of the most interesting SrtA inhibitors needs to be evaluated by in vivo models of infection, but we used a simple methodology in vitro, such as inhibition of biofilm formation, for a preliminary assessment of antivirulence properties of some of these novel inhibitors of SrtA. An interesting correlation between inhibition of SrtA and interference with the formation of staphylococcal biofilms has been observed in many cases especially at a concentration equal or more than IC50 determined as SrtA inhibitors.

Gli idrolati, o acque aromatiche, sono i prodotti dell'idrodistillazione di specie aromatiche. Possono essere ottenuti dall'intera pianta o parti di essa, allo stato fresco o in buona conservazione come fiori o, nel nostro caso, dai frutti del Genere Citrus. Le sue proprietà sono utilizzate nella produzione di alimenti, cosmetici e profumi, o in farmacia e aromaterapia. Essi sono comunemente considerati un sottoprodotto degli oli essenziali, ma in realtà, data la diversa composizione e la conseguente maggiore delicatezza, hanno un loro apprezzabile utilizzo, legato comunque al mantenimento delle proprietà delle piante da cui derivano. La produzione degli oli essenziali nell'industria agrumaria avviene tramite spremitura a freddo delle bucce e gli idrolati sono sostanza di scarto considerati un rifiuto da smaltire. Dal punto di vista chimico essi possono essere considerati delle soluzioni sature di oli essenziali (frazione volatile) in acqua. La composizione chimica di tali soluzioni è poco nota: si sa comunque che sono ricchi di composti prevalentemente ossigenati che oltre a dare un gradevole profumo, possono conferirgli numerose proprietà biologiche. Lo scopo di tale lavoro è quello di intraprendere uno studio di caratterizzazione LC-MS e GC-MS sull'idrolato dell’arancia di Ribera per identificare le sostanze presenti che potrebbero essere di interesse farmaceutico, cosmetico, ecc. e inoltre testarne la possibile attività biochimica e funzionale. Risultati positivi di questo studio potrebbero rappresentare un’opportunità per l’industria agrumaria, che vedrebbe la valorizzazione di un prodotto attualmente di scarto.

Medicinal mushrooms represent an unlimited source of polysaccharides with nutritional, antitumoral, antibacterial, and immune-stimulating properties. Traditional studies of epigeous higher Basidiomycetes have recently been joined by studies of hypogeous fungi and, in particular, of so-called desert truffles. With the aim to obtain novel agents against bacteria of clinical importance, we focused on the edible desert truffle mushrooms Tirmania pinoyi, Terfezia claveryi, and Picoa juniperi as sources of new antimicrobial agents. In particular, we investigated the in vitro antibacterial activity of acid-soluble protein extracts (aqueous extracts) of these 3 species against the Gram-positive human pathogenic reference strain Staphylococcus aureus ATCC 29213 and the Gram-negative strain Pseudomonas aeruginosa ATCC 15442. The acid-soluble protein extracts of T. pinoyi and T. claveryi showed minimum inhibitory concentrations of 50 μg/mL against tested pathogens. We believe that such preliminary results are promising to obtain a valuable antibiotic alternative to fight antibiotic-resistant pathogens.

Following our previous research on anti-inflammatory drugs (NSAIDs), we report on the design and synthesis of 4-(aryloyl)phenyl methyl sulfones. These substances were characterized for their capacity to inhibit cyclooxygenase (COX-1 and COX-2) isoenzymes. Molecular modeling studies showed that the methylsulfone group of these compounds was inserted deep in the pocket of the human COX-2 binding site, in an orientation that precludes hydrogen bonding with Arg120, Ser353, and Tyr355 through their oxygen atoms. The N-arylindole 33 was the most potent inhibitor of COX-2 and also the most selective (COX-1/COX-2 IC(50) ratio was 262). The indole derivative 33 was further tested in vivo for its anti-inflammatory activity in rats. This compound showed greater inhibitory activity than ibuprofen. Other compounds (20, 26, 9, and 30) showed strong activity against carrageenan-induced inflammation. The latter compounds showed a weak capacity to inhibit the proliferation of human cell lines K562, NCI-H460, and HT-29 in vitro.

New N-methyl and N-ethyl substitutions in the indazole nucleus are reported by reacting 3-(2-aminobenzamido)indazole and the appropriate trimethyl/triethyl orthobenzoate. Single crystal X-ray analysis confirms the N-ethylation position for the 3-(1-ethyl-1H-indazol-3-yl)-2-phenylquinazolin-4(3H)-one derivative 3f. Compounds 11a-d and 3a-d were tested to evaluate their antimicrobial, their antiproliferative activity and their COX inhibitory activities showing scarce or moderately antiproliferative activity and some inhibitory activity against COX-1 and COX-2.

Several new 2-(2-phenoxyacetamido)benzamides 17a-v, 21 and 22 were synthesized by stirring in pyridine the acid chlorides 16a-e and the appropriate5-R-4-R1-2-aminobenzamide 15a-e and initially evaluated in vitro for antiproliferative activity against the K562 (human chronic myelogenous leukemia) cell line. Some of synthesized compounds were evaluated for their in vitro antiproliferative activity against the full NCI tumor cell line panel derived from nine clinically isolated cancer types (leukemia, non-small cell lung, colon, CNS, melanoma, ovarian, renal, prostate and breast). The most active compounds caused an arrest of K562 cells in the G0-G1 phase of cell cycle and induction of apoptosis, which was mediated by caspase activation.

Gram-positive bacteria are a significant cause of nosocomial and community-acquired infections associated with diseases of high morbidity and mortality. Moreover, antibiotic resistance of important Gram-positive pathogens, such as Staphylococcus aureus, Streptococcus pneumoniae and Enterococcus faecalis is one of the major worldwide health problems. Over the last decade, many studies have focused on agents that target the virulence of important pathogens without killing or inhibiting their growth therefore imposing limited selective pressure to promote the development and spread of resistance mechanisms [1]enterococci and streptococci, sortase A plays a critical role in Gram-positive bacterial pathogenesis. It is thus considered a promising target for the development of new anti-infective drugs that aim to interfere with important Gram-positive virulence mechanisms, such as adhesion to host tissues, evasion of host defences, and biofilm formation. The additional properties of sortase A as an enzyme that is not required for Gram-positive bacterial growth or viability and is conveniently located on the cell membrane making it more accessible to inhibitor targeting, constitute additional reasons reinforcing the view that sortase A is an ideal target for anti-virulence drug development. Many inhibitors of sortase A have been identified to date using high-throughput or in silico screening of compound libraries (synthetic or natural. We synthesized two classes of molecules: curcumin analogues and N-phenyl-1H-pyrazole-4-carboxamides able to interfere with the bacterial adhesion, a fundamental step of Gram-positive pathogenesis [2].All derivatives obtained were tested for both the planktonic growth inhibition activity and the inhibition of biofilm formation against the following reference bacterial strains: S. aureus ATCC 6538, S. aureus ATCC 25923 and S. aureus ATCC 29213.

Increased attention has been focused on marine invertebrates as a source of bioactive molecules for biomedical applications. Many bioactive molecules are part of the innate immune system. Some more recently isolated compounds, mainly from the sea urchin and the sea cucumber, are antimicrobial peptides (AMPs) active against Gram-positive and Gram-negative bacteria, and fungi. In this review we described the most recent studies on AMPs isolated from echinoderms. AMPs are small peptides (< 10 kDa) with cationic charge and amphipathic structure. Recently, it was demonstrated that in the coelomocyte lysates of Paracentrotus lividus and Holothuria tubulosa, AMPs possess activity against staphylococcal and Pseudomonas aeruginosa biofilms. The data shows a great potential for application of AMPs in biotechnology for developing novel therapeutic agents that are either alternative or complementary to conventional antibiotic therapy to combat multi-resistant pathogens.

The rise of antibiotic-resistance as well as the deficiency of investments by pharmaceutical companies in the development of new antibiotics, have stimulated the investigation of alternative strategies to conventional antibiotics for counteracting the pathogens. A fundamental step of Gram positive pathogenesis is the bacterial adhesion to the host tissue involving a direct and a specific interaction between bacterial surface molecules and host ligands. Targeting the adhesion is a good strategy to design novel anti-infective drugs agents useful to interfere with the pathogenic process and with a virulence mechanism as biofilm formation. This review is focused on anti-virulence compounds which target bacterial surface molecules such as exposed proteins and theicoic acids and on their potential development as therapeutic agents alternative or complementary to conventional antibiotics in the contrast of Gram positive pathogens.The rise of antibiotic-resistance as well as the deficiency of investments by pharmaceutical companies in the development of new antibiotics, have stimulated the investigation of alternative strategies to conventional antibiotics for counteracting the pathogens. A fundamental step of Gram positive pathogenesis is the bacterial adhesion to the host tissue involving a direct and a specific interaction between bacterial surface molecules and host ligands. Targeting the adhesion is a good strategy to design novel anti-infective drugs agents useful to interfere with the pathogenic process and with a virulence mechanism as biofilm formation. This review is focused on anti-virulence compounds which target bacterial surface molecules such as exposed proteins and theicoic acids and on their potential development as therapeutic agents alternative or complementary to conventional antibiotics in the contrast of Gram positive pathogens.

The rise of antibiotic-resistance as well as the reduction of investments by pharmaceutical companies in the development of new antibiotics have stimulated the investigation for alternative strategies to conventional antibiotics. Many antimicrobial peptides show a high specificity for prokaryotes and a low toxicity for eukaryotic cells and, due to their mode of action the development of resistance is considered unlikely. We recently characterized an antimicrobial peptide that was called Paracentrin 1 from the 5-kDa peptide fraction from the coelomocyte cytosol of the Paracentrotus lividus. In this study, the chemically synthesized Paracentrin 1, was tested for its antimicrobial and antibiofilm properties against reference strains of Gram positive and Gram negative. The Paracentrin 1 was active against planktonic form of staphylococcal strains (reference and isolates) and Pseudomonas aeruginosa ATCC 15442 at concentrations ranging from 12.5 to 6.2 mg/ml. The Paracentrin 1 was able to inhibit biofilm formation of staphylococcal and Pseudomonas aeruginosa strains at concentrations ranging from 3.1 to 0.75 mg/ml. We consider the tested peptide as a good starting molecule for novel synthetic derivatives with improved pharmaceutical potential.

3,4-diphenylisoxazolo[3,4-d]pyridazin-7(6H)-one analogs were synthesized and tested for the antiproliferative activity. Study on the cell cycle alteration and on some cellular target (ATM, procaspase-2 proteins and H2AX histone) demonstrate the increase of the cell population in S phase and to induce cellular death by apoptosis by DNA damage with double strand breaks.

The reaction under reflux between 1-phenyl-3-R-5-methylaminopyrazoles and 2,5-hexanedione lead to 5,7:7,13-dimethanopyrazolo[3,4-b]pyrazolo[3′,4′:2,3]azepino[4,5-f]azocine derivatives 3b–g. These unusual molecules show the structural complexity of many biologically active natural products and are endowed with the chemical diversity that is required in drug discovery. The compounds 3b,e were reduced by hydrogen in the presence of Palladium on activated charcoal to give the dihydro derivatives 5b,e. Compounds 3b–f and 5b,e were selected by the NCI to evaluate their in vitro antiproliferative activity against 60 human cell lines derived from nine clinically isolated cancer types (leukaemia, lung, colon, melanoma, renal, ovarian, brain, breast, and prostate). The most active compound of this series, caused a block in G0–G1 phase of cell cycle. Analysis of pRb expression showed that this compound favours pRb dephosphorylation.

Essential oils from six different populations of Origanum vulgare subsp. hirtum were compared for their antibiofilm properties. The six essential oils (A to F) were characterized by a combination of gas chromatography with flame ionization detector and gas chromatography with mass spectrometer detector analyses. All oils showed weak activity against the planktonic form of a group of Staphylococcus aureus strains and against a Pseudomonas aeruginosa ATCC 15442 reference strain. The ability to inhibit biofilm formation was investigated at sub-MIC levels of 200, 100, and 50 m g/ml by staining sessile cells with safranin. Sample E showed the highest average effectiveness against all tested strains at 50 m g/ml and had inhibition percentages ranging from 30 to 52%. In the screening that used preformed biofilm from the reference strain P. aeruginosa, essential oils A through E were inactive at 200 m g/ml; F was active with a percentage of inhibition equal to 53.2%. Oregano essential oil can inhibit the formation of biofilms of various food pathogens and food spoilage organisms.Essential oils from six different populations of Origanum vulgare subsp. hirtum were compared for their antibiofilm properties. The six essential oils (A to F) were characterized by a combination of gas chromatography with flame ionization detector and gas chromatography with mass spectrometer detector analyses. All oils showed weak activity against the planktonic form of a group of Staphylococcus aureus strains and against a Pseudomonas aeruginosa ATCC 15442 reference strain. The ability to inhibit biofilm formation was investigated at sub-MIC levels of 200, 100, and 50 m g/ml by staining sessile cells with safranin. Sample E showed the highest average effectiveness against all tested strains at 50 m g/ml and had inhibition percentages ranging from 30 to 52%. In the screening that used preformed biofilm from the reference strain P. aeruginosa, essential oils A through E were inactive at 200 m g/ml; F was active with a percentage of inhibition equal to 53.2%. Oregano essential oil can inhibit the formation of biofilms of various food pathogens and food spoilage organisms.

The present study aims to investigate coelomocytes, immune mediators cells in the echinoderm Holothuria tubulosa, as an unusual source of antimicrobial and antibiofilm agents. The activity of the 5kDa peptide fraction of the cytosol from H. tubulosa coelomocytes (5-HCC) was tested against a reference group of Gram-negative and Gram-positive human pathogens. Minimal inhibitory concentrations (MICs) ranging from 125 to 500 mg/ml were determined against tested strains. The observed biological activity of 5-HCC could be due to two novel peptides, identified by capillary RP-HPLC/nESI-MS/MS, which present the common chemical-physical characteristics of antimicrobial peptides. Such peptides were chemically synthesized and their antimicrobial activity was tested. The synthetic peptides showed broad-spectrum activity at 12.5 mg/ml against the majority of the tested Gram-positive and Gram-negative strains, and they were also able to inhibit biofilm formation in a significant percentage at a concentration of 3.1 mg/ml against staphylococcal and Pseudomonas aeruginosa strains. The immune mediators in H. tubulosa are a source of novel antimicrobial peptides for the development of new agents against biofilm bacterial communities that are often intrinsically resistant to conventional antibiotics.

Several new benzamides 4a–q were synthesized by stirring in pyridine the acid chlorides 3a–q with the appropriate anthranilamide derivatives 2a–g. Some of the synthesized compounds were evaluated for their in vitro antiproliferative activity against a panel of 5 human cell lines (K562 human chronic myelogenous leukemia cells, MCF-7 breast cancer cells, HTC-116 and HT26 colon cancer cells and NCI H460 non-small cell lung cancer cells).

Several new 2-cinnamamido, 2-(3-phenylpropiolamido) and 2-(3-phenylpropanamido)benzamides were synthesized by stirring in pyridine the opportune acid chlorides with the appropriate anthranilamide derivatives. Some of the synthesized compounds were evaluated for their in vitro antiproliferative activity against a panel of 5 human cell lines (K562 human chronic myelogenous leukemia cells, MCF-7 breast cancer cells, HTC-116 and HT26 colon cancer cells and NCI H460 non-small cell lung cancer cells).

The immune mediators in echinoderms can be a potential source of novel antimicrobial peptides (AMPs) applied toward controlling pathogenic staphylococcal biofilms that are intrinsically resistant to conventional antibiotics. The peptide fraction5 kDa from the cytosol of coelomocytes of the sea urchin Paracentrotus lividus (5-CC) was tested against a group of Gram-positive and Gram-negative pathogen reference strains. The 5-CC of P. lividus was active against all planktonic-tested strains but also showed antibiofilm properties against staphylococcal strains. Additionally, we demonstrated the presence of three small peptides in the 5-CC belonging to segment 9-41 of a P. lividusβ-thymosin. The smallest of these peptides in particular, showed the common chemical-physical characteristics of AMPs. This novel AMP from β-thymosin has high potential activity as an antibiofilm agent, acting on slow-growing bacterial cells that exhibit a reduced susceptibility to conventional antibiotics and represent a reservoir for recurrent biofilm-associated infections.

Several new 4-diazopyrazole derivatives 6a – g and 9a – c were obtained by the reaction of 1-(R-substituted-phenyl)-3-(1,3-dimethyl-1 H -pyrazol-5-yl)ureas 5a – g and N -(1,3-dimethyl-1 H -pyrazol-5-yl)-2-(R-substituted-phenyl)acetamides 8a – c respectively with a sevenfold excess of nitrous acid in acetic acid solution. The compounds were assayed for their activity against the Staphylococcus aureus reference strains ATCC 25923, ATCC 29213 and ATCC 6538, as well as six veterinary strains. The best anti-staphylococcal profile was showed by [(R-substituted-phenyl)acetyl](4-diazonio-1,3-dimethyl-1 H -pyrazol-5-yl)azanides 9a , c . Compound 9c was also able at 3.1 μg mL −1 to inhibit of 45.7% the biofilm formation of the strains S. aureus ATCC 29213.

By reacting methylaminopyrazoles with hexane-2,5-dione in 1,4-dioxane in the presence of p-toluensulfonic acid, new derivatives containing the policyclic sistem 5,7:7,13-dimethanopyrazolo[3,4-b]pyrazolo[3'.4':2,3]azepino[4,5-f]azocine were obtained. The new synthesized compounds were tested preliminarly at 10 microM against five human cancer cell lines showing a range of inhibition of 20-62% against the most susceptible cell lines K562 and HCT116.

With the aim to face the threat of pathogen biofilms intrinsically resistant to conventional antibiotics, we focused on coelomocytes, the immune mediators in echinoderms, as source of novel antimicrobial peptides (AMPs). The proteic fraction

The red palm weevil (RPW), Rhynchophorus ferrugineus (Olivier, Coleoptera, Curculionidae) is the major pest of palm trees in the Mediterranean area. Damages are caused mainly by the endophytophagous larvae that consume the palm tender soft tissues making galleries and producing a wet “frass” with a characteristic fermented odour. The culturable bacterial community associated to the frass produced by RPW larvae inside Phoenix canariensis trees is mainly composed of facultative anaerobe gamma-proteobacteria that have their closest phylogenetic relatives in the gut microbiome of other endophytophagous and xilophagous coleopteran as bark beetles and other plant biomass-degrading insects (leaf-cutter ants). In RPW-bored galleries, these enterobacteria together with lactic acid bacteria, could be responsible for the synthesis of organic volatiles such as ethyl esters that are attractants for adults that deposit their eggs on the palm. The hypothesis that the frass-associated bacteria might be inoculated by the RPW larvae in order to obtain benefits for the insect (i.e. attraction of adults, plant polymers degradation, protection against fungal invasion, etc.) is under investigation. To this aim we started to characterize the bacterial community of the RPW larval gut by culture-based and culture-independent methods. Bacterial and DNA isolation was carried out from RPW larval guts after sterilization and dissection. About 107 CFU/gut were detected on Nutrient Agar plates and the phylogenetic analysis of 48 isolates is in progress. Cellulosolytic and hemicellulosolytic proteobacteria and actinobacteria were isolated from gut enrichment cultures on carboxymethyl cellulose and sterile filter paper but no cellulolytic activity was detected in the frass-associated bacterial community. Total gut DNA was used in 16S rRNA gene analysis by denaturing gradient gel electrophoresis and for the construction of a ribosomal gene library that is under screening.

The cinnamoyl anthranilamides represent a class of biological active substances of great importance in medicinal chemistry. Moreover, despite their wide range of biological activities, a review of the literature revealed that no anticancer activity is described for this kind of substances. Starting from the 2-cinnamamido-5-iodobenzamide, resulted able to inhibit the leukemic cell line K-562 proliferation with a percent of inhibition of 74% at 10M concentration, we undertake the following structural modifications on cinnamamidobenzamide skeleton: the introduction of various substituents both on the benzamido and the cinnamamido moieties, the substitution of olefinic bond with the ethane, ethyne, cyclopropane and phenyl groups. Synthesized compounds caused growth inhibition against many tumor cell lines at low micromolar and submicromolar concentrations against every tumor cell line investigated The best activity was obtained when the 5 position of the benzamido moiety was substituted with an iodine moiety. COMPARE analysis, effects on tubulin polymerization in cells and with purified tubulin, and effects on cell cycle distribution (G2-M phase block), including induction of apoptosis, indicate that these new antiproliferative compounds act as antitubulin agents. Preliminary biological data on 2-(3-phenylpropiolamido)benzamides, bearing an ethyne bond, showed a different mechanism of action considering that they caused a G0-G1 block of the cell cycle on K526 cell line.

Pyrazolylbenzotriazinones are endowed with structural analogy with the COX-2 selective inhibitor celecoxib. Considering that our research group has long been interested in the 3-pyrazolyl-substituted benzotriazinones as anti-inflammatory agents, six new pyrazolylbenzotriazinone derivatives 16a-c and 18a-c have been prepared by reacting the opportune ethyl 5-(2-aminobenzamido)-1-(pyridin-2-yl)-1H-pyrazole-4-carboxylate or 5-(2-aminobenzamido)-1-(pyridin-2-yl)-1H-pyrazole-4-carboxyic acid with sodium nitrite in glacial acetic acid. The biological studies revealed a good pharmacological profile for some pyrazolylbenzotriazinones and, in the case of the ethyl 5-(4-oxo-1,2,3-benzotriazin-3(4H)-yl)-1-pyridin-2-yl-1H-pyrazole-4-carboxylate 16a, a good COX-1/COX-2 selectivity. Molecular modeling studies confirmed the obtained biological results.

Some natural and synthetic related pyrrolomycins, a family of halogenated pyrrole antibiotics, showed anti-biofilm properties in vitro at low concentration (0.045μg/mL) against preformed staphylococcal biofilms. Moreover, considering the human cell toxicity, the selectivity indexes (ratio of cytotoxicity to antibiofilm activity) of some of them were very interesting. The present study aims to investigate if the pyrrolomycins could also prevent staphylococcal biofilm formation. The evaluation of S.aureus ATCC 25923 biofilm formation inhibition was conducted by safranin staining method. At tested concentrations of 0.18, 0.09, 0.045 μg/mL (concentrations much lower than MIC value determined on planktonic strain) the novel pyrrolomycin derivative IV resulted effective as biofilm inhibitor showing inhibition percentages ranging from 56.5 to 29% against S.aureus ATCC 25923. We are investigating if sortase A which is responsible for the anchoring of surface proteins to Gram positive cell wall, could be the rational target of pyrrolomycins. The surface proteins play pivotal roles in the adhesion to host’s tissues, and the evasion of host-immune responses, moreover they facilitate attachment on biological and abiotic surfaces in the first steps of biofilm formation. It has been observed that inhibition of sortase A by different chemicals resulted in decrease of virulence and staphylococcal biofilm formation. A molecular modeling study conducted by using a crystal structure of sortase A, recovered from protein data bank, and by studying docking properties of a known sortase A inhibitor and of pyrrolomycins, is in progress. In vitro experiments on sortase inhibition activity are needed to confirm the computational results

The pyrazolo[3,4-d]pyrimidine system shows a multitude of interesting pharmacological properties. Owing to the potential anti-inflammatory activity of 5-benzamido-pyrazolo[3,4-d]pyrimidin- 4-one derivatives and considering the easy synthesis of this class of compounds, a set of new 5- benzamido-1H-pyrazolo[3,4-d]pyrimidin-4-ones has been prepared in 42-80% yields by reacting 5- aminopyrazole-4(N-benzoyl)carbohydrazide derivatives and the opportune triethylorthoesters. Compounds 8a, b, 10a–d, and 11a, b revealed a superior inhibitory profile against COX-2, when compared to that of reference standards NS398 and indomethacin. Molecular modelling studies confirmed the obtained biological results.

The title compounds were easily synthesized by reacting the 4-aminopyrazole hydrochloride 2 and the substituted 2-nitrobenzoyl chlorides 3a-d. The obtained 2-nitrobenzamides 4a-d were methylated and then reduced to give the corresponding amines 6a-d. These were hydrolyzed then directly converted into 4,10-dihydro-1H-pyrazolo[3,4-c][1,5]benzodiazocine-5,11-diones 1a-d by the action of SOCl2 in benzene. These were tested for their COX inhibitory activity, showing an inhibitory profile against both COX-1 and COX-2, being slightly more selective against COX-2 with a percentage of inhibition, at the concentration of 10 μM, in the range 42.0 – 55.0.

Through the screening of a Streptomyces coelicolor genomic library, carried out in a histidinol phosphate phosphatase (HolPase) deficient strain, SCO5208 was identified as the last unknown gene involved in histidine biosynthesis. SCO5208 is a phosphatase, and it can restore the growth in minimal medium in this HolPase deficient strain when cloned in a high or low copy number vector. Moreover, it shares sequence homology with other HolPases recently identified in Actinobacteria. During this work a second phosphatase, SCO2771, sharing no homologies with SCO5208 and all so far described phosphatases was identified. It can complement HolPase activity mutation only at high copy number. Sequence analysis of SCO5208 and SCO2771, amplified from the HolPase mutant strain, revealed that SCO5208 shows a mutation in a conserved amino acid, whereas SCO2771 does not show any mutation. All these results show that S. coelicolor SCO5208, recently renamed hisN, is the HolPase involved in histidine biosynthesis.

Background: The recognition that chronic infections and infections associated with medical devices are biofilm related has been the impulse for investigating the antibiofilm properties of some diazopyrazoles biologically active as antimicrobials. Methods: The susceptibility of staphylococcal biofilms was determined at concentrations ranging from 25 to 1.5 µg/ml using crystal violet and methylthiazotetrazolium (MTT) staining. In the case of Candida albicans, we first assessed the anti-germ tube formation effect of 4-NO2 (compound 1c) and then we evaluated its antibiofilm activity at concentrations ranging from 100 to 6.2 µg/ml. Results: All the compounds were active against reference staphylococcal biofilm strains at the maximum tested concentration of 25 µg/ml; despite its remarkable activity as germ tube inhibitor, compound 1c showed weak activity against C. albicans biofilms at 100 µg/ml. Conclusion: 4-Diazopyrazoles represent an interesting source of antimicrobial compounds in the development of new antibiofilm agents.